Quenching-enhanced fluorescence titration protocol for accurate determination of free energy of membrane binding

Fluorescence spectroscopy is a convenient tool to examine peptide-membrane interactions at equilibrium ( 1 ) owing to the change in emission properties of many fluorophores (including tryptophan) during transfer from an aqueous environment into a lipid bilayer. In some cases (e.g. mechanosensitive c...

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Bibliographic Details
Published in:Analytical biochemistry 2007-03, Vol.362 (2), p.290-292
Main Authors: Posokhov, Yevgen O., Gottlieb, Philip A., Ladokhin, Alexey S.
Format: Article
Language:English
Subjects:
Animals
Melitten - chemistry
Melitten - metabolism
Membrane Lipids - chemistry
Membrane Lipids - metabolism
Protein Binding
Spectrometry, Fluorescence - methods
Thermodynamics
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Summary:Fluorescence spectroscopy is a convenient tool to examine peptide-membrane interactions at equilibrium ( 1 ) owing to the change in emission properties of many fluorophores (including tryptophan) during transfer from an aqueous environment into a lipid bilayer. In some cases (e.g. mechanosensitive channel blocker GsMTx4 described here), however, binding-associated emission changes are too small for reliable determination of the free energy of partitioning, ΔG. To enhance the spectroscopic response to binding we implemented the titration with lipid vesicles in the presence of aqueous ionic quencher iodide, which preferentially quenchers fluorescence of the free peptide in solution. We have verified the accuracy of this new titration protocol using the well-studied peptide melittin.
ISSN:0003-2697
1096-0309
DOI:10.1016/j.ab.2006.11.022